Median Fluorescence Intensity Research Articles

Desensitization therapy for elevated donor-specific antibody levels in haploidentical transplantation in Chilean patients

Allogeneic stem cell transplantation (ALOSCT) is curative for several hematological diseases. Unrelated donors and cord blood stem cells are valid options but haploidentical donors (HAPLO) have been considered the main source of stem cells in several countries, partly due to easy access and low cost of the transplantation process. Unfortunately, some patients have antibodies against the HLA epitopes from family haploidentical donors (donor-specific antibodies [DSA]) which are associated with engraftment failure and lethality. A few strategies exist to reduce or eliminate HLA antibodies that bind to these receptors. In this study, we present our experience with DSA desensitization by retrospectively examining a cohort from our hospital program. Between 2012 and 2023, we performed 243 ALOSCTs, of which 142 were from HAPLO and 56 were from unrelated donors. Nine patients (7%) had elevated DSA levels, most of which were female patients and mostly HAPLO. The median fluorescence intensity for these patients was 22,490 (19,000-28,560). Most of these high DSA patients (80%) received a desensitization procedure that involves plasmapheresis, rituximab, and immunoglobulin. The remaining 20% had severe infections during transplantation, and received rituximab monotherapy instead. The median dose of stem cells infused was 6.5 x 10^6 CD34/kg. Graft-versus-host disease (GVHD) preventative measures for all patients involved post-transplantation cyclophosphamide. Primary graft failure was observed in 45% of DSA elevated patients. For the remaining patients, median granulocyte and platelet engraftment were 14 d (12-16) and 16 d (13-18), respectively. Mortality in patients who did not receive engraftment was 100%. The incidence of mild, chronic GVHD was 15%. In conclusion, the desensitization of DSA in patients provided a 55% rate of engraftment and survival. However, a 45% rate of primary graft failure continued to pose a challenge in patients with DSA and required the development of improved strategies to reduce elevated transplant-related mortality.

Empirical sample-specific approaches to define HPV16 and HPV18 seropositivity in unvaccinated, young, sexually active women.

Given low seroconversion rates following human papillomavirus (HPV) infection, fixed external cutoffs may lead to errors in estimating HPV seroprevalence. We evaluated finite mixture modeling (FMM) and group-based trajectory modeling (GBTM) among unvaccinated, sexually active, HPV-exposed women to determine study-specific HPV16 and HPV18 seropositivity thresholds. We included 399 women (aged 18-24 years) enrolled in the HPV Infection and Transmission Among Couples Through Heterosexual Activity (HITCH) cohort study between 2005 and 2011 in Montreal, Canada. Participants' blood samples from up to six visits spanning 2 years were tested by multiplex serology for antibodies [median fluorescence intensity (MFI)] specific to bacterially expressed HPV16 and HPV18 L1 glutathione S-transferase fusion proteins. We applied FMM and GBTM to baseline and longitudinal antibody titer measurements, respectively, to define HPV type-specific seronegative and seropositive distributions. Study-specific thresholds were generated as five standard deviations above the mean seronegative antibody titers, mimicking cutoffs (HPV16: 422 MFI; HPV18: 394 MFI) derived from an external population of sexually inactive, HPV DNA-negative Korean women (aged 15-29 years). Agreement (kappa) of study-specific thresholds was evaluated against external cutoffs. Seroprevalence estimates using FMM (HPV16: 27.5%-43.2%; HPV18: 21.7%-49.5%) and GBTM (HPV16: 11.8%-11.8%; HPV18: 9.9%-13.4%) thresholds exceeded those of external cutoffs (HPV: 10.2%; HPV18: 9.7%). FMM thresholds showed slight-to-moderate agreement with external cutoffs (HPV16: 0.26%-0.46%; HPV18: 0.20%-0.56%), while GBTM thresholds exhibited high agreement (HPV16: 0.92%-0.92%; HPV18: 0.82%-0.99%). Kappa values suggest that GBTM, used for longitudinal serological data, and otherwise FMM, for cross-sectional data, are robust methods for determining the HPV serostatus without prior classification rules.IMPORTANCEWhile human papillomavirus (HPV) seropositivity has been employed as an epidemiologic determinant of the natural history of genital HPV infections, only a fraction of women incidentally infected with HPV respond by developing significant antibody levels. HPV seropositivity is often determined by a dichotomous fixed cutoff based on the seroreactivity of an external population of women presumed as seronegative, given the lack of evidence of HPV exposure. However, considering the variable nature of seroreactivity upon HPV infection, which arguably varies across populations, such externally defined cutoffs may lack specificity to the population of interest, causing inappropriate assessment of HPV seroprevalence and related epidemiologic uses of that information. This study demonstrates that finite mixture modeling (FMM) and group-based trajectory modeling (GBTM) can be used to independently estimate seroprevalence or serve as the basis for defining study-specific seropositivity thresholds without requiring prior subjective assumptions, consequently providing a more apt internally valid discrimination of seropositive from seronegative individuals.

Seroprevalence of sexually transmitted infections over 44 years - A cross-sectional study in Sweden.

Sexually transmitted infections (STIs) may cause substantial individual suffering and a large economic burden for society. This study examined the seroprevalence of Chlamydia trachomatis, Mycoplasma genitalium, herpes simplex virus (HSV) types 1 and 2, and several human papillomaviruses (HPV) in the Swedish population over time. The study population consisted of 30-year-old women attending maternity care, and 50year-old men and women attending health check-ups, from 1975 to 2018. Antibody status was determined by multiplex serology and quantified using median reporter fluorescence intensity (MFI). A total of 891 samples were analysed (519 from 30-year-old women, 186 from 50year-old women and 186 from 50year-old men). Of these, 41.5% showed seropositivity for Chlamydia trachomatis, 16.7% for Mycoplasma genitalium, 70.5% for HSV-1, 14.9% for HSV-2, 13.2% for high-risk HPV, and 8.3% for low-risk HPV. Seropositivity for Mycoplasma genitalium, HSV-1 and especially Chlamydia trachomatis decreased over time. There was a decrease over time in Chlamydia trachomatis seroprevalence, probably due to contact tracing, testing and early treatment; this might also have affected Mycoplasma genitalium seroprevalence. Despite the reduction, seroprevalences are still high, so continued and new efforts to reduce STI incidence are essential.

Abstract LB345: Correlating the blood immune repertoire and clinical features of del17p CLL patients

Abstract Introduction: Deletion in the short arm of chromosome 17 (del17p) is a strong, negative prognostic biomarker found in about eight percent of patients with untreated chronic lymphocytic leukemia (CLL). CLL patients with del17p exhibit resistance to therapy, accelerated disease progression, and global immunodeficiency. It is unclear the extent to which global immunodeficiency impacts disease course and therapy outcomes. The goal of this study was to document the blood immune repertoire of patients with untreated CLL who have del17p and associate with clinical features, time to first treatment (TFT), and overall survival (OS). Methods: Biobanked blood samples from treatment-naïve del17p CLL patients (n = 58) and age-matched controls (n = 5) were obtained. B cells and T cells were analyzed with comprehensive, cell type-specific flow cytometry panels and visualized with unbiased dimensionality reduction. Lab values were log-transformed and compared using t-tests and one-way ANOVAs. TFT was calculated from date of sample collection to date of first CLL-directed therapy or last known untreated date; OS was calculated from date of sample collection to death or last known alive status. Cox proportional hazards regression analyses (results reported as hazard ratio [HR] and 95% confidence interval [CI]) were employed to identify associations of lab values with OS and TFT. Results: Patients with del17p CLL had increased proportions of T regulatory cells (Tregs) among total CD4+ T cells compared to controls (CLL 11.1%, control 5.7%, p = 0.05). A higher proportion of CD8+ T cells was associated with longer TFT (HR 0.57, 95%CI 0.36-0.89); in contrast, higher proportions of TIGIT+ (HR 1.83, 95%CI 1.03-3.28) and TIM3+ conventional (non-Treg) CD4+ T cells were associated with shorter TFT (HR 1.70, 95%CI 1.02-2.84). A higher proportion of PD1+ CD8+ T cells was associated with shorter OS (HR 1.79, 95%CI 1.05-3.06), in addition to PD1+ conventional CD4+ T cell proportion (HR 2.21, 95%CI 1.06-4.60). CLL B cells expressed PD-1 and inducible T cell costimulator (ICOS) at significantly lower median fluorescence intensities (MFIs) than control B cells (p < 0.0001 for both). The CXCR5+ MFI (HR 1.0002, 95%CI 1.0001-1.003) and CD23+ MFI (HR 1.88, 95%CI 1.01-3.50) of CLL B cells were associated with shorter TFT. Discussion: In this cohort of treatment-naïve del17p CLL patients, we determined that patients with higher proportions of exhausted CD4+ and CD8+ T cell phenotypes had significantly shorter OS. Cytotoxic T cell proportion positively associated with TFT, whereas proportions of CD4+ T cell exhaustion phenotypes negatively associated with TFT. Higher CXCR5+ and CD23+ MFIs of CLL B cells also associated with shorter TFT. The preliminary findings from our study suggest that the blood immune repertoire has strong associations with clinically relevant outcomes such as time to first treatment and overall survival in patients with untreated del17p CLL. Citation Format: Ethan J. Katcher, Kim A. Gwin, Sameer A. Parikh, Kari G. Rabe, Petra K. Lothert, Preeti Trisal, Esteban Braggio, Susan L. Slager, Nicholas Strong, Daniel L. Van Dyke, Zhiquan Wang, C. Chris Huang, Anita K. Gandhi, Kay L. Medina, Neil E. Kay. Correlating the blood immune repertoire and clinical features of del17p CLL patients [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2024; Part 2 (Late-Breaking, Clinical Trial, and Invited Abstracts); 2024 Apr 5-10; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2024;84(7_Suppl):Abstract nr LB345.

Circulating immune cell phenotyping is potentially relevant for diabetic retinopathy risk assessment

BackgroundInflammation is believed to play a central role in the development of diabetes mellitus and is a common feature of type 2 diabetes mellitus (T2DM). However, the association with diabetic retinopathy (DR) remains a topic of debate. MethodsThis study employed two-sample bidirectional Mendelian randomization (MR) analyses to establish causal associations between immune cell characteristics and DR. Using publicly available GWAS genetic data, we investigated the causal relationship between 731 immune cell characteristics and the risk of DR. A total of four types of immune features, including relative cell (RC), absolute cell (AC), median fluorescence intensities (MFI), and morphological parameters (MP), were included. Sensitivity analysis was conducted to assess the robustness, heterogeneity, and potential horizontal pleiotropy of the results. ResultsThirty-five immune cell phenotypes were correlated with the risk of developing DR among four immune traits (MFI, RC, AC, and MP), and DR resulted in altered expression of twenty-six immune cells. ConclusionWe have demonstrated a strong correlation between immune cell traits and DR using a genetic approach. This finding offers valuable insights for early DR prevention and future clinical research and treatment.

The enigmatic interplay of immune cells and abnormal spermatozoa through Mendelian randomization.

Abnormal spermatozoa significantly impact reproductive health, affecting fertility rates, potentially prolonging conception time, and increasing the risk of miscarriages. This study employs Mendelian randomization to explore their potential link with immune cells, aiming to reveal their potential causal association and wider implications for reproductive health. We conducted forward and reverse Mendelian randomization analyses to explore the potential causal connection between 731 immune cell signatures and abnormal spermatozoa. Using publicly available genetic data, we investigated various immune signatures such as median fluorescence intensities (MFI), relative cell (RC), absolute cell (AC), and morphological parameters (MP). Robustness was ensured through comprehensive sensitivity analyses assessing consistency, heterogeneity, and potential horizontal pleiotropy. The MR study produced a statistically significant p-value of.0000684, Bonferroni-corrected for the 731 exposures. The Mendelian randomization analysis revealed strong indications of a reciprocal relationship between immune cell pathways and sperm integrity. When examining immune cell exposure, a potential causal link with abnormal sperm was observed in 35 different types of immune cells. Conversely, the reverse Mendelian randomization results indicated that abnormal sperm might causally affect 39 types of immune cells. These outcomes suggest a potential mutual influence between alterations in immune cell functionality and the quality of spermatozoa. This study highlights the close link between immune responses and sperm development, suggesting implications for reproductive health and immune therapies. Further research may offer crucial insights into male fertility and immune disorders.

ShinyMBA: a novel R shiny application for quality control of the multiplex bead assay for serosurveillance studies

The multiplex bead assay (MBA) based on Luminex xMAP technology can be used as a tool to measure seroprevalence as part of population immunity evaluations to multiple antigens in large-scale serosurveys. However, multiplexing several antigens presents challenges for quality control (QC) assessments of the data because multiple parameters must be evaluated for each antigen. MBA QC parameters include monitoring bead counts and median fluorescence intensity (MFI) for each antigen in plate wells, and performance of assay controls included on each plate. Analyzing these large datasets to identify plates failing QC standards presents challenges for many laboratories. We developed a novel R Shiny application, shinyMBA, to expedite the MBA QC processes and reduce the risk of user error. The app allows users to rapidly merge multi-plate assay outputs to evaluate bead count, MFI, and performance of assay controls using statistical process control charts for all antigen targets simultaneously. The utility of the shinyMBA application and its various outputs are demonstrated using data from 32 synthetic xPONENT files with 3 multiplex antigens and two population serosurveillance studies that evaluated 1200 and 3871 samples, respectively, for 20 multiplexed antigens. The shinyMBA open-source code is available for download and modification at https://github.com/CDCgov/shinyMBA. Incorporation of shinyMBA into Luminex serosurveillance workflows can vastly improve the speed and accuracy of QC processes.

Wiskott-Aldrich syndrome protein expression in female WAS carriers: A flow cytometry study from North India.

Wiskott-Aldrich syndrome (WAS) is a rare X-linked inborn error of immunity characterized by microthrombocytopenia, infections, eczema, and increased predisposition to develop autoimmunity and malignancy. Flow cytometric assay for determining WAS protein (WASp) is a rapid and cost-effective tool for detecting patients. However, very few studies described WASp expression in female carriers. Most WAS carriers are clinically asymptomatic. Active screening of female family members helps identify female carriers, distinguish de novo mutations, and to select appropriate donor prior to curative stem cell transplantation. This study was undertaken to evaluate the diagnostic capability of flow cytometry-based WASp expression in peripheral blood cells to identify carriers and compare WASp expression in different blood cell lineages. Female patients, heterozygous for WAS gene, were enrolled in this study conducted at Pediatric Allergy Immunology Unit, Advanced Pediatric Centre, Post Graduate Institute of Medical Education and Research, Chandigarh, India. Flow cytometric assessment of WASp expression in lymphocytes, monocytes, and neutrophils was carried out and compared with healthy control and affected patients. The results were expressed in delta (Δ) median fluorescence intensity (MFI) as well as stain index (SI), which is the ratio of ΔMFI of patient and ΔMFI of control. Thirteen mothers and two sisters of genetically confirmed WAS patients were enrolled in the study. All enrolled females were clinically asymptomatic and did not have microthrombocytopenia. Low WASp expression (SI<1) was seen in lymphocytes and monocytes in 10 (66.6%) carriers. Females with variants in proximal exons (exons 1 and 2) were found to have lesser expression than those with distal (exons 3-12) variants. Flow cytometry is a rapid, easily available, cost-effective tool for WASp estimation. Lymphocytes followed by monocytes are the best cell lineages for WASp estimation in carrier females. However, genetic testing remains the gold standard, as carrier females with variants in distal exons may have normal WASp expression.

Abstract 5184: Biomarker results from PEVENAZA, a randomized phase 2 study of venetoclax and azacitidine +/- pevonedistat in newly diagnosed AML patients unfit for intensive chemotherapy: Increased efficacy in a subset of patients with IDH1/2 mutations and other observations

Abstract Background: Many newly diagnosed patients with AML are ineligible for intensive chemotherapy due to pre-existing comorbidities and older age. VIALE-A, a phase-3 trial with venetoclax (ven) and azacitidine (aza) established this combination as a new standard of care (SOC) for this patient population. The addition of pevonedistat (pevo) to this combination was evaluated in the same patient population in the current trial (NCT04266795). Here we present results of biomarker analyses from baseline and on-treatment bone marrow samples. Methods: Molecular mutational analysis was performed using a next generation sequencing (NGS) panel on baseline bone marrow samples. Combining this data with cytogenetics allowed centralized assignment of risk category based on ELN2017 guidelines. Longitudinal bone marrow aspirate samples were also examined by multi-color flow cytometry for expression of select surface antigen expression and to identify AML leukemic stem cells (LSCs) from the blast population. Results: Eleven genes were selected for analyses based on prevalence and scientific interest, including DNMT3A, ASXL1, TET2, FLT3, and IDH1/2. In contrast to other genes and ELN risk categories, we observed a statistically significant increase (p-value 0.013) in CR+CRi rates for IDH1/2 mutant patients when treated with pevo + ven + aza (95%, n=21) compared to ven+aza control arm (62%, n=21). In addition, we evaluated the prevalence of CD33, CD123 and CLEC12a, targets of current AML developmental therapeutics, in both the blast and LSC populations at baseline. The median (n=120 pts) % blast cells expressing CD33, CD123, or CLEC12a were 89.8%, 63.2%, and 66.0%, respectively. The median (n=78 pts) % LSCs expressing CD33, CD123, or CLEC12a were 48.2%, 90.9%, and 32.0%, respectively. Importantly, we did not see any significant difference in median fluorescence intensity (MFI) or % expression in any of the three proteins after treatment with ven and aza (+/- pevo). Finally, no strong associations were observed between expression levels and either mutations or ELN risk categories. Conclusions: Analyses of biomarker samples as part of the PEVENAZA trial resulted in several insights for not only the pevonedistat program but also for other AML therapeutics in development. The observation that newly diagnosed patients with IDH1/2 mutations may benefit from the addition of pevo to ven+aza is certainly hypothesis-generating but would require further follow-up with higher patient numbers. In addition, baseline CD33, CD123, and CLEC12a expression levels were in line with published studies. Here we have also shown they remain similar after SOC treatment (ven+aza) supporting the development of agents that target these proteins in both first and second line settings for AML. Citation Format: Radha Ramesh, Xiang Fang, Shuli Li, Sharon Friedlander, Farhad Sedarati, Lionel Adès, Nicholas Short, Cristina Papayannidis, Tammie Yeh. Biomarker results from PEVENAZA, a randomized phase 2 study of venetoclax and azacitidine +/- pevonedistat in newly diagnosed AML patients unfit for intensive chemotherapy: Increased efficacy in a subset of patients with IDH1/2 mutations and other observations [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2024; Part 1 (Regular Abstracts); 2024 Apr 5-10; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2024;84(6_Suppl):Abstract nr 5184.

Platelet reactivity in young children undergoing congenital heart disease surgery: a NITRIC randomized clinical trial substudy

IntroductionThe international NITRIC trial studied the hypothesis that nitric oxide (NO) applied into the cardiopulmonary bypass (CPB) oxygenator in infants would improve recovery after heart surgery. In a substudy, we evaluated the effect of NO applied into the CPB oxygenator on the (re)activity of platelets measured as fibrinogen binding (platelet aggregation) and P-selectin expression (platelet degranulation) in young children.MethodsPlatelet activity (without agonist exposure) and reactivity (after stimulation by an agonist) was studied in a single center substudy of the NITRIC trial, a multicenter, randomized trial that studied administration of 20 parts per million (ppm) NO during CPB in children younger than 2 years. Blood was collected at 4 time points (T1- T4); before CPB, after CPB start, before and after weaning. Flow cytometry-based platelet activity and reactivity in the presence of 5 agonists was tested. Differences on P-selectin expression and fibrinogen binding (median fluorescence intensity (MFI)) were analyzed with mixed effect modelling (MEM).ResultsBlood samples were obtained in 22 patients allocated to NO and 20 controls. Platelet counts dropped after T1 due to the hemodilution of blood in all patients (p < 0.001). Beta coefficients for NO allocation derived from the MEM models on fibrinogen binding and P-selectin expression were small (standardized beta coefficients on fibrinogen binding were 0.07[0.03, 0.11] and on P-selectin expression 0.05[0.03, 0.08]) and non-significant. CPB duration did not affect platelet reactivity (standardized beta coefficients 0.09[0.02, 0.12] with p > 0.27) in any of the MEMs.Conclusion20 ppm NO administration in the sweep gas of the CPB oxygenator did not affect platelet reactivity in young children undergoing heart surgery. Interestingly, duration of CPB exposure also did not have an effect on platelet (re)activity.Trial registrationANZCTR, ACTRN12617000821392. Registered 5 June 2017, https://anzctr.org.au/

Causal role of immune cells in psoriasis: a Mendelian randomization analysis.

A growing body of evidence has shown that immune cells are linked to psoriasis. It is, however, still unclear if these associations reflect a relationship of cause and effect. We employed a two-sample Mendelian randomization (MR)-based study to elucidate the probable causative connection between immune cells and psoriasis. Summary information for psoriasis (Ncase = 5,427, Ncontrol = 479,171) was obtained from the European Bioinformatics Institute. Summarized statistical information on 731 immune cell features, including morphological parameters (MP; n = 32), relative cell number (n = 192), median fluorescence intensity (MFI) of surface antigens (n = 389), and absolute cell number (n = 118), was obtained from the genome-wide association studies (GWAS) catalog. The research consisted of forward MR analysis, in which immune cell traits were used as the exposure factor, and psoriasis was the outcome, as well as reverse MR analysis, in which psoriasis was used as the exposure factor, and immune cell traits were the outcome. We ran numerous sensitivity analyses to ascertain the study results for robustness, heterogeneity, and potential multiple-biological effects. This research determined a probable causative connection between immune cells and psoriasis. In particular, we identified 36 distinct types of immune cells that are potentially causally linked to psoriasis. Our findings indicate strong causal correlations between 36 immunological phenotypes and psoriasis, thus, directing future clinical trials.

Impaired immune responses in the airways are associated with poor outcome in critically ill COVID-19 patients

Mounting evidence indicates that an individual's humoral adaptive immune response plays a critical role in the setting of SARS-CoV-2 infection, and that the efficiency of the response correlates with disease severity. The relationship between the adaptive immune dynamics in the lower airways with those in the systemic circulation, and how these relate to an individual's clinical response to SARS-CoV-2 infection, are less understood and are the focus of this study. Here, we investigate the adaptive immune response to SARS-CoV-2 in paired samples from the lower airways and blood from 27 critically ill patients during the first wave of the pandemic (median time from symptom onset to intubation 11 days). Measurements included clinical outcomes (mortality), broncheoalveolar lavage (BAL) and blood specimen antibody levels, and BAL viral load. While there was heterogeneity in the levels of the SARS-CoV-2 specific antibodies, we unexpectedly found that some BAL specimens displayed higher levels than in paired concurrent plasma samples, despite the known dilutional effects common in BAL samples. We found that survivors had higher levels of anti-Spike, anti-Spike-NTD, and anti-Spike-RBD IgG antibodies in their BAL (6684 [258–13 148]versus15 899 [8958–22 629], 5336 [256–10 343]versus13 494 [8028–19 414], and 1620 [199–6637]versus8466 [5144–16 157] median fluorescent intensity units, for deceasedversussurvivors, respectively, p&lt;0.05), while there was no such association with antibody levels in the systemic circulation. Thus, our data highlight the critical role of local adaptive immunity in the airways as a key defense mechanism against primary SARS-CoV-2 infection.

Unraveling the causal role of immune cells in gastrointestinal tract cancers: insights from a Mendelian randomization study.

Despite early attempts, the relationship between immune characteristics and gastrointestinal tract cancers remains incompletely elucidated. Hence, rigorous and further investigations in this domain hold significant clinical relevance for the development of novel potential immunotherapeutic targets. We conducted a two-sample Mendelian randomization (MR) analysis using the tools available in the "TwoSampleMR" R package. The GWAS data for these 731 immune traits were sourced from the GWAS Catalog database. Concurrently, data on gastrointestinal tract cancers, encompassing malignant tumors in the esophagus, stomach, small intestine, colon, and rectum, were extracted from the FinnGen database. The immune traits subjected to MR analysis predominantly fall into four categories: median fluorescence intensities (MFI), relative cell (RC), absolute cell (AC), and morphological parameters (MP). To ensure the reliability of our findings, sensitivity analyses were implemented to address robustness, account for heterogeneity, and alleviate the impact of horizontal pleiotropy. A total of 78 immune traits causally linked to gastrointestinal tract cancers were identified, encompassing esophageal cancer (12 traits), gastric cancer (13 traits), small intestine cancer (22 traits), colon cancer (12 traits), and rectal cancer (19 traits). Additionally, 60 immune traits were recognized as protective factors associated with gastrointestinal tract cancers, distributed across esophageal cancer (14 traits), gastric cancer (16 traits), small intestine cancer (7 traits), colon cancer (14 traits), and rectal cancer (9 traits). Furthermore, it was observed that seven immune traits are causally related to gastrointestinal tract cancers in at least two locations. These traits include "CCR2 on CD14- CD16+ monocyte," "CD19 on IgD+ CD38-," "CD19 on IgD+ CD38- naive," "CD25hi CD45RA+ CD4 not Treg AC," "CD27 on unsw mem," "CD28 on CD39+ activated Treg," and "CD45 on CD4+." This study elucidates a causal link between immune cells and gastrointestinal tract cancers at various sites through genetic investigation. The findings of this research open up new perspectives and resources for exploring tumor prevention strategies and immunotherapeutic targets.

Gene association analysis to determine the causal relationship between immune cells and juvenile idiopathic arthritis

BackgroundJuvenile idiopathic arthritis (JIA) is a type of chronic childhood arthritis with complex pathogenesis. Immunological studies have shown that JIA is an acquired self-inflammatory disease, involving a variety of immune cells, and it is also affected by genetic and environmental susceptibility. However, the precise causative relationship between the phenotype of immune cells and JIA remains unclear to date. The objective of our study is to approach this inquiry from a genetic perspective, employing a method of genetic association analysis to ascertain the causal relationship between immune phenotypes and the onset of JIA.MethodsIn this study, a two-sample Mendelian randomization (MR) analysis was used to select single nucleotide polymorphisms (SNPs) significantly associated with immune cells as instrumental variables to analyze the bidirectional causal relationship between 731 immune cells and JIA. There were four types of immune features (median fluorescence intensity (MFI), relative cellular (RC), absolute cellular (AC), and morphological parameters (MP)). Finally, the heterogeneity and horizontal reproducibility of the results were verified by sensitivity analysis, which ensured more robust results.ResultsWe found that CD3 on CM CD8br was causally associated with JIA at the level of 0.05 significant difference (95% CI = 0.630 ~ 0.847, P = 3.33 × 10−5, PFDR = 0.024). At the significance level of 0.20, two immunophenotypes were causally associated with JIA, namely: HLA DR on CD14+ CD16- monocyte (95% CI = 0.633 ~ 0.884, P = 6.83 × 10–4, PFDR = 0.16) and HLA DR on CD14+ monocyte (95% CI = 0.627 ~ 0.882, P = 6.9 × 10−4, PFDR = 0.16).ConclusionOur study assessed the causal effect of immune cells on JIA from a genetic perspective. These findings emphasize the complex and important role of immune cells in the pathogenesis of JIA and lay a foundation for further study of the pathogenesis of JIA.

Causal role of immune cells in obstructive sleep apnea hypopnea syndrome: Mendelian randomization study

BACKGROUND Despite being one of the most prevalent sleep disorders, obstructive sleep apnea hypoventilation syndrome (OSAHS) has limited information on its immunologic foundation. The immunological underpinnings of certain major psychiatric diseases have been uncovered in recent years thanks to the extensive use of genome-wide association studies (GWAS) and genotyping techniques using high-density genetic markers (e.g. , SNP or CNVs). But this tactic hasn't yet been applied to OSAHS. Using a Mendelian randomization analysis, we analyzed the causal link between immune cells and the illness in order to comprehend the immunological bases of OSAHS. AIM To investigate the immune cells' association with OSAHS via genetic methods, guiding future clinical research. METHODS A comprehensive two-sample mendelian randomization study was conducted to investigate the causal relationship between immune cell characteristics and OSAHS. Summary statistics for each immune cell feature were obtained from the GWAS catalog. Information on 731 immune cell properties, such as morphologic parameters, median fluorescence intensity, absolute cellular, and relative cellular, was compiled using publicly available genetic databases. The results' robustness, heterogeneity, and horizontal pleiotropy were confirmed using extensive sensitivity examination. RESULTS Following false discovery rate (FDR) correction, no statistically significant effect of OSAHS on immunophenotypes was observed. However, two lymphocyte subsets were found to have a significant association with the risk of OSAHS: Basophil %CD33dim HLA DR- CD66b- (OR = 1.03, 95%CI = 1.01-1.03, P &lt; 0.001); CD38 on IgD+ CD24- B cell (OR = 1.04, 95%CI = 1.02-1.04, P = 0.019). CONCLUSION This study shows a strong link between immune cells and OSAHS through a gene approach, thus offering direction for potential future medical research.

Association of immune cell traits with Parkinson's disease: a Mendelian randomization study.

Immunity and neuroinflammation play crucial roles in the pathogenesis of Parkinson's disease (PD). Nonetheless, prior investigations into the correlation between immune inflammation and PD have produced varying results. Identifying specific immune cell phenotypes that are truly associated with PD is challenging, and the causal relationship between immune cells and PD remains elusive. This study conducted a comprehensive two-sample Mendelian randomization (MR) analysis, employing five distinct analytical approaches, to clarify the causal connection between immune cell characteristics and the risk of PD. Utilizing GWAS data, we investigated the causal relationship between 731 immune cell traits and PD. These immune cell phenotypes encompass absolute cell (AC) counts, median fluorescence intensity (MFI), and relative cell (RC) counts for B cells, cDCs, mature stage T cells, monocytes, myeloid cells, TBNK (T cells, B cells, and natural killer cells), and Tregs, as well as the logistic parameter (MP) for cDCs and TBNK. The inverse variance weighted (IVW) analysis indicated that Myeloid DCs (p = 0.004), HVEM expression on CD45RA- CD4+ T cells (p = 0.007), CD62L- CD86+ Myeloid DCs (p = 0.015), and HLA DR expression on monocytes (p = 0.019) were associated with a reduced risk of PD. CD14+ CD16+ monocytes (p = 0.005), HLA DR+ NK cells within CD3- lymphocytes (p = 0.023), and CD28 expression on activated & secreting Tregs (p = 0.032) were associated with an increased risk of PD. This study establishes a causal link between immune cell phenotype and the pathogenesis of PD, identifying several specific immune cell characteristics associated with PD. This could inspire researchers to delve into the pathogenesis of PD at the cellular subtype level, and aid in the identification of potential pharmacological protein targets for PD.

Genetic insights into the relationship between immune cell characteristics and ischemic stroke: A bidirectional Mendelian randomization study.

Ischemic stroke, a major contributor to global disability and mortality, is underpinned by intricate pathophysiological mechanisms, notably neuroinflammation and immune cell dynamics. Prior research has identified a nuanced and often paradoxical link between immune cell phenotypes and ischemic stroke susceptibility. The aim of this study was to elucidate the potential causal links between the median fluorescence intensity (MFI) and morphological parameters (MP) of 731 immune cell types and ischemic stroke risk. By analyzing extensive genetic datasets, we conducted comprehensive Mendelian randomization (MR) analyses to discern the genetic correlations between diverse immune cell attributes (MFI and MP) and ischemic stroke risk. Our study identified key immune cell signatures linked to ischemic stroke risk. Both B cells and T cells, among other immune cell types, have a bidirectional influence on stroke risk. Notably, the regulatory T-cell phenotype demonstrates significant neuroprotective properties, with all odds ratio (OR) values and confidence intervals (CIs) being less than 1. Furthermore, CD39 phenotype immune cells, particularly CD39+ CD8+ T cells (inverse variance weighting [IVW] OR 0.92, 95% CI 0.87-0.97; p = 0.002) and CD39+ activated CD4 regulatory T cells (IVW OR 0.93, 95% CI 0.90-0.97; p < 0.001), show notable neuroprotection against ischemic stroke. This investigation provides new genetic insights into the interplay between various immune cells and ischemic stroke, underscoring the complex role of immune processes in stroke pathogenesis. These findings lay a foundation for future research, which may confirm and expand upon these links, potentially leading to innovative immune-targeted therapies for stroke prevention and management.

Application of Flow Cytometry Using Advanced Chromatin Analyses for Assessing Changes in Sperm Structure and DNA Integrity in a Porcine Model.

Chromatin status is critical for sperm fertility and reflects spermatogenic success. We tested a multivariate approach for studying pig sperm chromatin structure to capture its complexity with a set of quick and simple techniques, going beyond the usual assessment of DNA damage. Sperm doses from 36 boars (3 ejaculates/boar) were stored at 17 °C and analyzed on days 0 and 11. Analyses were: CASA (motility) and flow cytometry to assess sperm functionality and chromatin structure by SCSA (%DFI, DNA fragmentation; %HDS, chromatin maturity), monobromobimane (mBBr, tiol status/disulfide bridges between protamines), chromomycin A3 (CMA3, protamination), and 8-hydroxy-2'-deoxyguanosine (8-oxo-dG, DNA oxidative damage). Data were analyzed using linear models for the effects of boar and storage, correlations, and multivariate analysis as hierarchical clustering and principal component analysis (PCA). Storage reduced sperm quality parameters, mainly motility, with no critical oxidative stress increases, while chromatin status worsened slightly (%DFI and 8-oxo-dG increased while mBBr MFI-median fluorescence intensity-and disulfide bridge levels decreased). Boar significantly affected most chromatin variables except CMA3; storage also affected most variables except %HDS. At day 0, sperm chromatin variables clustered closely, except for CMA3, and %HDS and 8-oxo-dG correlated with many variables (notably, mBBr). After storage, the relation between %HDS and 8-oxo-dG remained, but correlations among other variables disappeared, and mBBr variables clustered separately. The PCA suggested a considerable influence of mBBr on sample variance, especially regarding storage, with SCSA and 8-oxo-dG affecting between-sample variability. Overall, CMA3 was the least informative, in contrast with results in other species. The combination of DNA fragmentation, DNA oxidation, chromatin compaction, and tiol status seems a good candidate for obtaining a complete picture of pig sperm nucleus status. It raises many questions for future molecular studies and deserves further research to establish its usefulness as a fertility predictor in multivariate models. The usefulness of CMA3 should be clarified.

Abstract WMP120: MicroRNAs as a Therapeutic Target to Reduce Microglial Activation After Post-Stroke Social Isolation

Introduction: Loneliness and social isolation (SI) are associated with increased risk of stroke and other vascular disorders, along with all-cause mortality. However, it is unknown what mechanisms mediate how social factors affect stroke recovery. Methods: To investigate the negative effects of post-stroke SI on brain miRNA profiles, aged (18-20 months) C57BL/6 male mice were utilized. Three days following a 60-minute transient right middle cerebral artery occlusion (MCAO), mice were randomly assigned to pair housing (PH) or single housing (SI). At post-stroke timepoints (specifically day 3 after the stroke), the infarct was equal between groups, reducing the possibility of differential effects resulting from different infarct sizes. At two-time intervals (post-stroke SI D4 and D27), temporal miRNA profiling of the ipsilateral hemisphere was performed. Results: Distinct miRNA candidates were significantly altered in post-stroke SI, within the brain across both acute and chronic time points (n=4/grp, FDR adjusted *p&lt;0.05). MiR-10a-5p and miR-10b-5p were observed to have crucial nodes in the pool of miRNAs that interacted with the largest group of miRNAs for post-stroke at SI D4 and D27, respectively. KEGG pathway analysis showed 4 days of isolation resulted in the enrichment of pathways related to microglial activation and 27 days of isolation lead to the activation of neuronal-specific pathways that regulate cognition and motivation (FDR adjusted *p&lt;0.05). At post-stroke SI D4, significant microglial activation was observed in independent validation cohorts as assessed by the median fluorescence intensity (MFI) of purinergic receptor P2Y12 (P2RY12), in CD45 int CD11b + P2RY12 + cells in the brain. MFI of P2RY12 was significantly downregulated in post-stroke SI mice at D4 (n=7-8/grp, *p&lt;0.05) compared to PH mice. Conclusions: These findings validate our hypothesis that post-stroke SI exacerbates microglia activation and leads to differential expression of miRNAs involved in regulating microglial pathways. Taken together, utilizing miRNA-based therapies to restore miRNA-profiles in the brain may be able to alleviate the detrimental impact of post-stroke SI on chronic depressive-like phenotypes.

Causal role of immune cells in generalized anxiety disorder: Mendelian randomization study.

Generalized anxiety disorder (GAD) is a prevalent emotional disorder that has received relatively little attention regarding its immunological basis. Recent years have seen the widespread use of high-density genetic markers such as SNPs or CNVs for genotyping, as well as the advancement of genome-wide association studies (GWAS) technologies, which have facilitated the understanding of immunological mechanisms underlying several major psychiatric disorders. Despite these advancements, the immunological basis of GAD remains poorly understood. In light of this, we aimed to explore the causal relationship between immune cells and the disease through a Mendelian randomization study. The summary information for GAD (Ncase=4,666, Ncontrol=337,577) was obtained from the FinnGen dataset. Summary statistics for the characterization of 731 immune cells, including morphological parameters (MP=32), median fluorescence intensity (MFI=389), absolute cells (AC=118), and relative cells (RC=192), were derived from the GWAS catalog. The study involved both forward MR analysis, with immune cell traits as the exposure and GAD as the outcome, and reverse MR analysis, with GAD as the exposure and immune cell traits as the outcome. We performed extensive sensitivity analyses to confirm the robustness, heterogeneity, and potential multi-biological effects of the study results. Also, to control for false positive results during multiple hypothesis testing, we adopted a false discovery rate (FDR) to control for statistical bias due to multiple comparisons. After FDR correction, GAD had no statistically significant effect on immunophenotypes. Several phenotypes with unadjusted low P-values are worth mentioning, including decreased PB/PC levels on B cells(β=-0.289, 95%CI=0.044~0.194, P=0.002), reduced PB/PC AC in GAD patients (β=-0.270, 95% CI=0.77~0.92, P=0.000), and diminished PB/PC on lymphocytes (β=-0.315, 95% CI=0.77~0.93, P=0.001). GAD also exerted a causal effect on CD27 on IgD-CD38br (β=-0.155,95%CI=0.78~0.94,P=0.002), CD20-%B cell (β= -0.105,95% CI=0.77~0.94, P=0.002), IgD-CD38br%lymphocyte(β=-0.305, 95%CI=0.79~0.95, P=0.002), FSC-A level on granulocytes (β=0.200, 95%CI=0.75~0.91, P=8.35×10-5), and CD4RA on TD CD4+(β=-0.150, 95% CI=0.82~1.02, P=0.099). Furthermore, Two lymphocyte subsets were identified to be significantly associated with GAD risk: CD24+ CD27+ B cell (OR=1.066,95%CI=1.04~1.10,P=1.237×10-5),CD28+CD4+T cell (OR=0.927, 95%CI=0.89~0.96, P=8.085×10-5). The study has shown the close association between immune cells and GAD through genetic methods, thereby offering direction for future clinical research.